Department of Veterinary Medicine, Università degli Studi di Milano, Via dell'Università 6, 26900, Lodi, Italy.
Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Epidemiology, Südufer 10, 17493, Greifswald-Insel Riems, Germany.
Parasit Vectors. 2021 Jan 13;14(1):47. doi: 10.1186/s13071-020-04557-6.
Neospora caninum, a coccidian protozoan, represents an important cause of bovine abortion. Available N. caninum strains show considerable variation in vitro and in vivo, including different virulence in cattle. To which extent sexual recombination, which is possible in the intestines of domestic dogs and closely related carnivores as definitive hosts, contributes to this variation is not clear yet.
Aborted bovine foetuses were collected between 2015 and early 2019 from Italian Holstein Friesian dairy herds suffering from reproductive problems. A total of 198 samples were collected from 165 intensive farms located in Lombardy, northern Italy. N. caninum samples were subjected to multilocus-microsatellite genotyping using ten previously established microsatellite markers. In addition to our own data, those from a recent study providing data on five markers from other northern Italian regions were included and analysed.
Of the 55 samples finally subjected to genotyping, 35 were typed at all or 9 out of 10 loci and their individual multilocus-microsatellite genotype (MLMG) determined. Linear regression revealed a statistically significant association between the spatial distance of the sampling sites with the genetic distance of N. caninum MLMGs (P < 0.001). Including data from this and a previous North Italian study into eBURST analysis revealed that several of N. caninum MLMGs from northern Italy separate into four groups; most of the samples from Lombardy clustered in one of these groups. Principle component analysis revealed similar clusters and confirmed MLMG groups identified by eBURST. Variations observed between MLMGs were not equally distributed over all loci, but predominantly observed in MS7, MS6A, or MS10.
Our findings confirm the concept of local N. caninum subpopulations. The geographic distance of sampling was associated with the genetic distance as determined by microsatellite typing. Results suggest that multi-parental recombination in N. caninum is a rare event, but does not exclude uniparental mating. More comprehensive studies on microsatellites in N. caninum and related species like Toxoplasma gondii should be undertaken, not only to improve genotyping capabilities, but also to understand possible functions of these regions in the genomes of these parasites.
刚地弓形虫是一种肉孢子虫原生动物，是牛流产的重要原因。现有的刚地弓形虫株在体外和体内表现出相当大的变异，包括在牛中的不同毒力。在肠道中作为终宿主的家养犬和近亲肉食动物中发生的性重组在多大程度上导致了这种变异尚不清楚。
2015 年至 2019 年初，从意大利荷斯坦弗里斯奶牛养殖场因生殖问题而流产的牛胎儿中收集了 198 个样本。这些样本来自意大利北部伦巴第地区的 165 个集约化农场。使用先前建立的 10 个微卫星标记对刚地弓形虫样本进行了多位点微卫星基因分型。除了我们自己的数据外，还包括了来自最近一项研究的数据，该研究提供了来自意大利北部其他地区的 5 个标记的数据，并进行了分析。
最终进行基因分型的 55 个样本中，有 35 个样本在 10 个位点中的 9 个或全部位点上进行了分型，并确定了它们各自的多位点微卫星基因型（MLMG）。线性回归显示，采样点的空间距离与刚地弓形虫 MLMG 的遗传距离之间存在统计学显著关联（P<0.001）。将本研究和意大利北部以前的一项研究的数据纳入 eBURST 分析显示，意大利北部的一些刚地弓形虫 MLMG 分为四个组；伦巴第的大多数样本聚集在其中一个组中。主成分分析显示出相似的聚类，并证实了 eBURST 识别的 MLMG 组。在 MLMG 之间观察到的变异并不是在所有位点上均匀分布的，而是主要在 MS7、MS6A 或 MS10 中观察到。
我们的研究结果证实了牛刚地弓形虫亚种群的存在。采样的地理距离与微卫星分型确定的遗传距离相关。结果表明，刚地弓形虫中的多亲本重组是一种罕见事件，但不能排除单亲交配。应该对刚地弓形虫和相关物种（如弓形虫）中的微卫星进行更全面的研究，不仅要提高基因分型能力，还要了解这些区域在这些寄生虫基因组中的可能功能。
